Fragile X Syndrome (FXS) is the most common inherited form of mental retardation. Almost all FXS is associated with nonexpressing null alleles of the X-linked FMR1 gene. A mouse model, fmr1-tm1Cgr, exists in which an internal exon of the murine FMR1 gene has been modified. The behavioral and cognitive deficits observed for fmr1-tm1Cgr mice are quite mild, absent, or unexpected compared to what is usually seen in human FXS. In particular, humans with FXS show a pronounced deficit in the ability to hold sequences of information in working memory. We have now compared the fmr1-tm1Cgr mice with their wild type littermates in tests of working memory, including retention of sequences of information as well as recall over variable delays. The fmr1-tm1Cgr mice performed as well as their wild type littermates, contrary to what would be expected from the human FXS phenotype. Significantly, we have generated several lines of evidence, subsequently confirmed in other laboratories, indicating that the fmr1-tm1Cgr allele is not a molecular null. We have found that: 1) normal levels of FMR1 mRNA are produced from fmr1-tm1Cgr's intact promoter, 2) at least one of the many spliced fmr1-tm1Cgr mRNAs has a full length open reading frame, and 3) the protein produced by the fmr1-tm1Cgr allele is observed at reduced levels in the neurons of the fmr1-tm1Cgr mouse brain. These findings raise the possibility that low levels of FMRP present in fmr1-tm1Cgr mice throughout development may be responsible for the surprisingly mild phenotype observed in these animals. We propose to create a true molecular null mouse model of FXS. We will then compare the cognitive and behavioral phenotype of the null animal with the extant fmr1-tm1Cgr mouse. If the null mice have a substantial cognitive impairment compared to the fmr1-tm1Cgr mice, this would suggest that a low level of FMRP could potentially have dramatic protective or curative effects in FXS. Among many other experiments, a molecular null animal will also allow the creation of conditional transgenic animals to determine the effects of turning FMRP on and off at various times in development. Therefore, the production of an FMR1 null mouse should provide the basis for significant new research into the nature of FXS and its treatment.